Evidence that plasma membrane electrical potential is required for vesicular stomatitis virus infection of MDCK cells: A study using fluorescence measurements through polycarbonate supports

Summary We used fluorescence microscopy of Madin-Darby Canine Kidney (MDCK) cells grown on polycarbonate filters to study a possible link between plasma membrane electrical potential (_pm) and infectivity of vesicular stomatitis virus (VSV). Complete substitution of K⁺ for extracellular Na⁺blocks VSV infection of MDCK cells as well as baby hamster kidney (BHK) cells. When we independently perfused the apical and basal-lateral surfaces of high resistance monolayers, high K⁺ inhibited VSV infection of MDCK cells only when applied to the basal-lateral side; high K⁺ applied apically had no effect on VSV infection. This morphological specificity correlates with a large decrease in _pm of MDCK cells when high K⁺ buffer is perfused across the basal-lateral surface. Depolarization of the plasma membrane by 130 mm basal K⁺ causes a sustained increase of cytosol pH in MDCK cells from 7.3 to 7.5 as reported by the fluorescent dye BCECF. Depolarization also causes a transient increase of cytosol Ca²⁺ from 70 to 300 nm as reported by the dye Fura-2. Neither increase could explain the block of VSV infectivity by plasma membrane depolarization. One alternative hypothesis is that _pm facilitates membrane translocation of viral macromolecules as previously described for colicins, mitochondrial import proteins, and proteins secreted by Escherichia coli.We thank Kenneth Spring for many helpful discussions concerning fluorescence digitized imaging systems, James Russell for his collaboration in the design of our imaging system, Herbert Chase for suggestions on dye loading into MDCK cells, and Manfred Schubert and George Harmison for providing expertise on VSV.     

Frequent Unanticipated Alleles of lethal giant larvae in Drosophila Second Chromosome Stocks

Forty years ago, a high frequency of lethal giant larvae (lgl) alleles in wild populations of Drosophila melanogaster was reported. This locus has been intensively studied for its roles in epithelial polarity, asymmetric neural divisions, and restriction of tissue proliferation. Here, we identify a high frequency of lgl alleles in the Bloomington second chromosome deficiency kit and the University of California at Los Angeles Bruinfly FRT40A-lethal P collection. These unrecognized aberrations confound the use of these workhorse collections for phenotypic screening or genetic mapping. In addition, we determined that independent alleles of insensitive, reported to affect asymmetric cell divisions during sensory organ development, carry lgl deletions that are responsible for the observed phenotypes. Taken together, these results encourage the routine testing of second chromosome stocks for second-site alleles of lgl.     

The Use of the Ex Vivo Chandler Loop Apparatus to Assess the Biocompatibility of Modified Polymeric Blood Conduits

The foreign body reaction occurs when a synthetic surface is introduced to the body. It is characterized by adsorption of blood proteins and the subsequent attachment and activation of platelets, monocyte/macrophage adhesion, and inflammatory cell signaling events, leading to post-procedural complications. The Chandler Loop Apparatus is an experimental system that allows researchers to study the molecular and cellular interactions that occur when large volumes of blood are perfused over polymeric conduits. To that end, this apparatus has been used as an ex vivo model allowing the assessment of the anti-inflammatory properties of various polymer surface modifications. Our laboratory has shown that blood conduits, covalently modified via photoactivation chemistry with recombinant CD47, can confer biocompatibility to polymeric surfaces. Appending CD47 to polymeric surfaces could be an effective means to promote the efficacy of polymeric blood conduits. Herein is the methodology detailing the photoactivation chemistry used to append recombinant CD47 to clinically relevant polymeric blood conduits and the use of the Chandler Loop as an ex vivo experimental model to examine blood interactions with the CD47 modified and control conduits.     

Learning and Long-Term Retention of Large-Scale Artificial Languages

Recovering discrete words from continuous speech is one of the first challenges facing language learners. Infants and adults can make use of the statistical structure of utterances to learn the forms of words from unsegmented input, suggesting that this ability may be useful for bootstrapping language-specific cues to segmentation. It is unknown, however, whether performance shown in small-scale laboratory demonstrations of “statistical learning” can scale up to allow learning of the lexicons of natural languages, which are orders of magnitude larger. Artificial language experiments with adults can be used to test whether the mechanisms of statistical learning are in principle scalable to larger lexicons. We report data from a large-scale learning experiment that demonstrates that adults can learn words from unsegmented input in much larger languages than previously documented and that they retain the words they learn for years. These results suggest that statistical word segmentation could be scalable to the challenges of lexical acquisition in natural language learning.     

Continuous Flow Routes toward Designer Metal Nanocatalysts

Mono‐ and multimetallic nanoparticles (NPs) have diverse and tunable physicochemical properties that arise from their compositions as well as crystallite size and shape. The ability to control precisely the composition and structure of NPs through synthesis is central to achieving state‐of‐the‐art designer metal NPs for use as catalysts and electrocatalysts. However, a major limitation to the use of designer metal NPs as catalysts is the ability to scale their syntheses while maintaining structural precision. To address this challenge, continuous flow routes to metal NPs involving the use of droplet microreactors are being developed, providing the synthetic versatility necessary to achieve known and completely new nanostructures. This progress report outlines how the chemistry and process parameters of droplet microreactors can be used to achieve high performing nanocatalysts through control of NP composition, size, shape, and architecture and outlines directions toward previously unimaginable nanostructures.     

Engineering Local Coordination Environments of Atomically Dispersed and Heteroatom‐Coordinated Single Metal Site Electrocatalysts for Clean Energy‐Conversion

Carbon‐based heteroatom‐coordinated single‐atom catalysts (SACs) are promising candidates for energy‐related electrocatalysts because of their low‐cost, tunable catalytic activity/selectivity, and relatively homogeneous morphologies. Unique interactions between single metal sites and their surrounding coordination environments play a significant role in modulating the electronic structure of the metal centers, leading to unusual scaling relationships, new reaction mechanisms, and improved catalytic performance. This review summarizes recent advancements in engineering of the local coordination environment of SACs for improved electrocatalytic performance for several crucial energy‐convention electrochemical reactions: oxygen reduction reaction, hydrogen evolution reaction, oxygen evolution reaction, CO₂ reduction reaction, and nitrogen reduction reaction. Various engineering strategies including heteroatom‐doping, changing the location of SACs on their support, introducing external ligands, and constructing dual metal sites are comprehensively discussed. The controllable synthetic methods and the activity enhancement mechanism of state‐of‐the‐art SACs are also highlighted. Recent achievements in the electronic modification of SACs will provide an understanding of the structure–activity relationship for the rational design of advanced electrocatalysts.     

Uncovering Atomic‐Scale Stability and Reactivity in Engineered Zinc Oxide Electrocatalysts for Controllable Syngas Production

In this study, scalable, flame spray synthesis is utilized to develop defective ZnO nanomaterials for the concurrent generation of H₂ and CO during electrochemical CO₂ reduction reactions (CO₂RR). The designed ZnO achieves an H₂/CO ratio of ≈1 with a large current density (j) of 40 mA cm^?2 during long‐term continuous reaction at a cell voltage of 2.6 V. Through in situ atomic pair distribution function analysis, the remarkable stability of these ZnO structures is explored, addressing the knowledge gap in understanding the dynamics of oxide catalysts during CO₂RR. Through optimization of synthesis conditions, ZnO facets are modulated which are shown to affect reaction selectivity, in agreement with theoretical calculations. These findings and insights on synthetic manipulation of active sites in defective metal‐oxides can be used as guidelines to develop active catalysts for syngas production for renewable power‐to‐X to generate a range of fuels and chemicals.     

Plant carbon allocation drives turnover of old soil organic matter in permafrost tundra soils

Carbon cycle feedbacks from permafrost ecosystems are expected to accelerate global climate change. Shifts in vegetation productivity and composition in permafrost regions could influence soil organic carbon (SOC) turnover rates via rhizosphere (root zone) priming effects (RPEs), but these processes are not currently accounted for in model predictions. We use a radiocarbon (bomb‐¹⁴C) approach to test for RPEs in two Arctic tall shrubs, alder (Alnus viridis (Chaix) DC.) and birch (Betula glandulosa Michx.), and in ericaceous heath tundra vegetation. We compare surface CO₂ efflux rates and ¹⁴C content between intact vegetation and plots in which below‐ground allocation of recent photosynthate was prevented by trenching and removal of above‐ground biomass. We show, for the first time, that recent photosynthate drives mineralization of older (>50 years old) SOC under birch shrubs and ericaceous heath tundra. By contrast, we find no evidence of RPEs in soils under alder. This is the first direct evidence from permafrost systems that vegetation influences SOC turnover through below‐ground C allocation. The vulnerability of SOC to decomposition in permafrost systems may therefore be directly linked to vegetation change, such that expansion of birch shrubs across the Arctic could increase decomposition of older SOC. Our results suggest that carbon cycle models that do not include RPEs risk underestimating the carbon cycle feedbacks associated with changing conditions in tundra regions.     

Unexpected side effects in biocrust after treating non‐native plants using carbon addition

Carbon addition has been proposed as an alternative to herbicide and manual removal methods to treat non‐native plants and reduce non‐target effects of treatments (e.g. impacts on native plants; surface disturbance). On Mojave Desert pavement and biocrust substrates after experimental soil disturbance and carbon addition (1,263 g C/m² as sucrose), we observed declines in lichens and moss cover in sucrose‐treated plots. To further explore this unforeseen potential side effect of using carbon addition as a non‐native plant treatment, we conducted biocrust surveys 5 and 7 years after treatments, sampled surface soils to observe if treatments additionally affected soil filamentous cyanobacteria, and conducted laboratory trials testing the effects of different levels of sucrose on cyanobacteria and desert mosses. Sucrose addition to biocrust plots reduced lichen and moss cover by 33–78% and species richness by 40–80%. Sucrose reduced biocrust cover in biocrust plots to levels similarly detected in pavement plots (<1%). While cyanobacteria in the field did not appear to be affected by sucrose, laboratory tests showed negative effects of sucrose on both cyanobacteria and mosses. Cyanobacteria declined by 41% 1 month after exposure to 5.4 g C/m² equivalent solutions. We detected injury to photosynthesis in mosses after 96 hour exposure to 79–316 g C/m² equivalent solutions. Caution is warranted when using carbon addition, at least in the form and concentration of sucrose, as a treatment for reducing non‐native plants on sites where conserving biocrust is a goal.